Method for the protection of loci susceptible to the growth of undesired microorganisms



United States Patent '1 3,275,503 METHOD FOR THE PROTECTION OF LOCI SUS- CEPTIBLE TO THE GROWTH OF UNDESIRED NIICROORGANISMS Lawrence F.1Marnett and Ralph]. 'Tenney, Kansas City,

Mo., and Jerome B. Thompson, Cumberland, Md., as-

signors to Q]. Patterson Company, Kansas City, M0.

N0. Drawing. Filed Aug..26, 1964,.Ser. No. 392,297 12 Claims. (Cl. 167-22) The present invention relates to an improved method for the protection of loci susceptible ,to the growth of undesired microorganisms, such as bacteria and fungi, for instance, molds,mildews and yeasts,.against such growth.

The term undesired microorganisms is employed herein to signify microorganisms whose growth is undesired in the particular, locus tobe'protected although they might be considered desired microorganisms in other situations.

As is well known, the growth and proliferation of many bacteria molds, mildews and yeasts is undesired in many locations because of deterioration of materials concerned, toxicological hazards, diesease causing factors and similar considerations. For example, foods and feedstutfs may be subjectto deterioration or spoilage because of growth of bacteria, molds or yeasts therein or thereon or materials, such as textiles,.1eather,an d the like, may be subject to deterioration both as to mechanical properties and/ or asto appearance-because of growth of mildews or molds thereon. Also, certainskin diseases, such as those .of the type of athlet'e?s foot are known to be caused or instigated by growth of types of fungi.

It is an object of the invention to provide an improved means for protecting -lo ci susceptible to the growth of undesired microorganisms against such growth. According to the invention it was unexpectedly found that certain acylated productsof a-hydroxy acids are very effective inhibitors of a wide spectrum of selective microorganisms such ,as molds, mildews, yeast-s and bacteria. The acyl lactylic-acid products employed according to the invention are of the-following general formula valence of the cation, m isan'integer offrom'l to n, x is a number froml-to. about 6, preferably 1 to about 3, and .each of A and Bare hydrogen or methyl, preferably one Patented Sept. 27, 1966 of A is hydrogen and the other is methyl which means that the product is an acyllactylic product of the formula CH3 [RCO(OCHCO)xO]Y(OH)n-m The acylatedproducts of a-hydroxy acids concerned according to the invention are produced by methods analogous to that described in 'U.S. Patent 2,733,252 directed to the production of salts of fatty acid esters of lactylic acids using the appropriate fatty acid and u-hydroxy carboxylic acid. The ,productsrin general aremixtures and therefore the values for x and m generally are average values. Nevertheless, separations such as-on a chromatographic column can be eifected. Also generally even when salts are prepared, in practical scale processes, some free acids are present in the product so that the product normally is slightly acid. Furthermore, the presence of free fatty acids, which has not been esterified with 41-11} droxy acid, or their'salts along with the acylated products of w-hydroxy acids of the =above'formula is not deleterious and-at most only exerts a diluting-action as thefatty acids andsalts concerned in some instances -possess activity against microorganisms although not as marked or as of broad a spectra as the acylated products according to the invention. In some instances the quantity of free fatty acids or their salts which may be present is such that the composition taken as a whole, that is, free fatty acid or its salt plus the acyl-ated products gives a product in which the value of x is less than 1, for example, about 0.3. Such compositions can be obtained by simple admixture of the free fatty acids or their salts with the acylated products or by preparation of the acylated products using less than 1 mol of a-hydroxy acid per mol of fatty acid.

The procedure employed for the production of the acylated o't-hydroxy carboxylic acids involves heating of the fatty acid concerned with the a-hYdI'OXY acid or'condensation products thereof in proportions adapted to provide the desired value for x in the presence of more than a catalytic amount of a basic reacting alkali metalor alkaline earth metal compound. When free acid products (Y='H) are desired they are easily prepared byacidiiication of the products with a dilute mineral acid, such as hydrochloric acid. Such free acid products can also be used in the preparation of various salts by reaction with the appropriate base ormetal compound. In defining the products concerned according to the invention, for example, as acyl 2 lactylic products, it is intended to indicate that such product contains an average of about 2 lactyl groups (x=2).

The following Tables 1 and 2 illustrate the procedures which can be employed and the properties of the products produced in tabular form.

TABLE 1.-WEIGHTS 0F REACTANTSAND REACTION CONDITIONS USED IN THE SYNTHESIS OF SALTS OFAC-YIi. 2 LACTYLIO ACID Grams Grams Reaction Reaction Acid Used Fatty Grams Acid Alkali Time, Hours Temper- Acid v ature, C.

Lactic Carbonate Caprylie acid 187. 5 243.3 (Ca) 65. 0 6 .200 Caprle acid 171. 2 179.10 (Ca) 49.8 1 /6 200 Laurie acid 186.0 167.6 (Ca) 46. 4 1% 200 Caprylic acid- 187. 5 234.3 (N a) s 68: 9 6% 200 Capric Acid- 2,06. 8 216. 2 (Na) 63: 6' 6 200 Laurie acid 1, 201. 8 1', 228.6 (N3) 318; 0 1% 200 lelycpnc T MO n V Capric acid 95.3 s4. 2 20-5 2 200 'd ox'yiso- C600] A uty l Capric acid 68. 0' 71. 5 19175 6 200 Lactic Ca(QH)i Undeeanoic acid.-- '20. 0 19. 8 4.1 3

\TABLE 2.ANALYSIS OF PREPARATIONS OF SALTS OF high?! efieFtiVefleSS than when x is higher, for example t ACYL-2 LACTYLIO ACIDS especially in the case of the capryl lactylates.

The pH at which the compounds employed according to I pH (27 Percent Acid Ester i suspensign) C8 or Na value Value to invention are most effective depends upon the partlcu f lar microorganism concerned. While they show excellent Calcium caDWlyl-Zlactylate. 4.5 6.6 101 240 effectweness at values 'i i ,Caicium capryl-21actylate 4.3 5 143 172 ample, 8.5, against some microorganisms theineffective- 4 8 7 9 60 181 ness against the broadest spectrum of microorganisms is !gaeium laury1l-%l2a1ctylalte 2. 1 g 132 in the acid range, particularly at pH values below 6.5. p 5 0 ium capryyacty ate l 190 Sodium cap WM 1a ct ylate 4. 6 0 134 178 10 Excellent rnhlbltion of yeasts other than Saccharomyces s or num mum- 2 2lacitylaltet g; 3.3 1:3 138 cerevlslae is also obtained as indicated by the following a cium eapry g yco a e. 1 182 1 Calcium caprym amyism results obtained with calcium eapryl 2 lactylate at a 0.1% butyrate 4.6 6.6 145 74 level.

I The efiectrveness of the acylated a-hydroxy carboxyllc Control Zgfigfiii acid products according to the invention in inhibiting V lactylfite growth of bacteria, molds and yeasts wasevaluated by g dispersing each of such compounds at experimental levels g g i fi t fl I T 0 in heated culture media composed of p 521%; '1: 3

E Indus. 0

1 Phytone 'grams 10 Dextrose do 10 The improved effectiveness of the compounds accord- Agar d0 16 ing to the invention over that of known inhibitors is illus- 1 Water liter 1 trated by the following table.

TABLE 4.LEVEL 0.1% OF THE CULTURE MEDIA Sorbic Sodium; Sodium Calcium 3 Control Acid Benzoate Propionate Oapryl-2 f V Lactylate 0 0 0 0 Aspergzllus mger 0 Trichophyton mentagrophytea 0 0 Saccharomuces cereuisiae 0 0 Asperaillus terreus 0 ++L- For comparison, culture media containing no additive The acylated products of puahydroxy carboxylic acids I Were included in a The culture d was 40 employed according to the invention as undesired micro- 3 poured into the required 100 mm. by 15 mm. petrrdishes. organism growth i hi i can be i d i h h These individual culture plates wereinoculated from cul- 1 or products to be Protected or such materials may tures of the selected microorganisms and incubated for be impregnated or mated therewith. In many instances were inspected and the growth of the microorganisms repers for goods to be Protected againstigmwth of micro corded. of 1 f h d Table 3 below gives the'results obtained with a 0.1% meamsms exam! wrappers ease Pm concentration of the followin com ounds:

g p ment and preventlon of diseases-of, for example, the skin a P 'Y Y l caused by microorganisms, the inhibitor eompoundscan OaCu2L=ca1cium undecanoyl'z lactylatc dusting powders, ointments, salves or solutions. In genggg gf ifi g i i q f Tgg fii eral, the variousmodes' of application of the. inhibitor l g z i lactysllate compounds according to the invention involves: close asm sociation with the locus to be protected against growth NaC 2L=sodium launyl-Z lactylate CaCm2G=oa1Gium capryla glycolate of undesired microorganisms. Of course, whenprotec- CaC 2-a-O=calcium capryl-2-a-oxyisobutyrate tion of food products and contact or association with liv- TABLE 3.-LEVEL 0.1% 013 THE CULTURE MEDIA Control 030821, CaCm2L cacnzr. 03cm NaC 2L NaCm2L' NaouzL CaCw2G CSCmQaO s: n z 0 0 o v '0 0 0 I 0 0 o affizziifliiiiiiili. 0 0 0 o o o 0 o Rhizome fliaric ns +-lI- Penicillium zptmsum 0 0 0 V0 Aspemillus nia r 0 0 0 gicliOphjIflOn 'mentagrpphutea g (6 d6 46 g 3 g 8 (1 mn C88 CBTEUISMZG--- Asil eru illu tarrem 0 Tests carried out with other salts, such as, for example, 70 ing bodies is concerned, it is understood that the cation 7 zinc and aluminum salts, and with the free acids (Y=H) of the inhibitor compound is not such as to render such showed an effectiveness similar to that of the alkali and compound toxic during its intended use. alkaline earth metal salts, indicating that the acyl laotylate The practical effectiveness of the compounds employed anion is responsible for the effectiveness. In addition,'it according to the invention is illustrated by the following was found that those compounds wherein x is 1 to 3 have 7 5 examples.

hours at 32 C. At the end of this period, the plates it is merely necessary for example,,io impregnate Wm? with the inhibitor compounds ofthe invention. For treat- I CaCmZLicalcium capryl'z lactylate 50 be applied topically to the areas afiected in the form of i A pan.

Example 1 .T he inhibition of the growth of Staphylococcus aureus in custard pie filling A premix containing all the dry ingredients fora custard pie filling was prepared according to the following formula:

Parts Sugar 7 42.5 Non-fat milk solids 21.2 Corn starch 13.3 Dried whole eggs 21.0 Salt 0.7 Vanilla 1.3

Seven sample pie fillings were prepared from the premix. The inhibitors, calcium capryl-2 lactylate and sorbic acid, were added to suspensions of the ingredients in Water. Since pH frequently effects the performance of inhibitors in foods, the two inhibitors were compared at three different pH levels. Citric acid, a commonly used .food additive which also occurs naturally in foods, was

used as the acidulant and was added to the fillings at levels shown. The fillings were cooked over steam until the consistency was similar to custard. The seven filling variables were as follows:

FORMULATION OF EXPERIMENTAL PIE FILLINGS USED TO STUDY STAPHYLOOOCC'LS AUREUS INHIBITION 'gram of inoculated filling. The counts were:

EFFECTIVENESS OF CALCIUM CAPRYL2 LACIYLATE AND SORBIC ACID ON THE INHIBITION OF STAPHYLO OOCC'USAUREUS INCUSTARD PIE FILLINGS Filling Identification Filling Bacteria per pH gram Filling 1. No additive. 6. 4 235,000, 000 .2. 0.2% Calcium Capry1-2 Lactylate 6. 1 5, 000, 000 .3. 0.2% SorbicAeid 6:1 22, 000, 000 "1. 0.2% Calcium Capryl-2 Lactylate 5.1 5.02% Calcium Capryl-Z Lactylate. 4. 85 Less than 100 6. 0.2% Sorbic Acid 5.15 00 7. ).2% Serbia Acid 4. 8 4, 700

Example 2.-Inhibz'tion of mildew development of canvas by impregnating canvas with calcium capryl-2 lactylate Canvas was cut into eight sections, six inches by six inches, soaked and .washed in a detergent, well rinsed and dried. Seven sections were dipped in ethanol solutions of 20%, 10%, 5%, 2.5% and 1% calcium capryl-2 ;lactylate and 5% and 1% sodium benzoate by weight.

.The excess solution wassqueezed out and the canvas sec- ..tions allowed to dry over night. The eight sections were buried under about one inch of soil contained in a large Water was added periodically to keep soil and canvas moist and to insure the development of mildew by the soil micro-organisms.

After thirty days the canvas was removed from the soil, carefully washed and examined. The condition of the cloth, according to treatment, was rated as follows:

First: (best condition) 20% calcium capryl-2 lactylate Second: 10% calcium capryl-2 lactylate Third: 5% calcium capryl2 lactylate Fourth: 2.5% calcium capryl-2 lactylate Fifth: 1% calcium capryl-2 lactylate and 5% benzoate Sixth: 1% sodium benzoate and the untreated control 63 mm. strips of each canvas section were tested on the Elmendorf Tearing Tester, a standard instrument used by the paper industry. The instruments graduations are used as an index of resistance to tearing. The results were:

Canvas treatment: Elmendorf units 20% calcium capryl-2 lactylate Over 100. 10% calcium capryl- 2 lactylate Over 100. 5% calcium capryl-2 lactylate Over 100. 2.5% calcium capryl-2 lactylate 55. 1% calcium capryl-2 lactylate 35. 5%sodium benzoate 19. 1% sodium benzoate 12. Control, no treatment 8.

The high resistance to tearing shown by the canvas treated with the capryl lactylate indicated this treatment reduced rotting of the fabric.

The capryl lactylate is more effective than sodium benzoate.

Example 3.Prevention of mold development on packaged cheese with the use of calcium capryl-2 lactylate Br-own'kraft paper was cut into three and a half inch squares. Four squares were dipped in ethanol; four in 10% by weight calcium capryl-2 lactylate in ethanol;

.four in 10% by weight suspension of sorbic acid .in

ethanol; four were not treated. The sheets were dried ,untilno odor ,of ethanol .was detected in the paper. These paper squares and twelve slices of American .cheese, .containing no preservative, .were ,spread on .a

table and exposed to air borne mold spores for thirty minutes. Each set of four squares was placed between, above and below, three slices of cheese. These sets were sealed in polyethylene bags and stored at room temperature.

The packages were inspected periodically and the dates on which the first mold growth and mold covereage appeared was recorded. Mold was first noted after four days in the packages containing the untreated paper and the paper treated only with ethanol. In seven more days these two packages were covered with mold. No mold developed in the packagecontaining papers treated with calcium capryl-2 lactylate and the sorbic acid.

Example 4.-In'hibition of mold growth on partially baked dinner rolls by application of calcium capryl-2 lactylate Partially baked dinner rolls were preparedaccording to commercial formulas and procedures. Forty-eight rolls were prepared and divided into four groups of twelve rolls. Each group was treatedas follows:

7 (1) Control, no treatment .conditions. Periodically the rolls were inspected and the time in days for the appearance of the first mold colony,

multiple colonies and complete mold coverage Was recorded. The results were:

STUDIES OF THE MOLD INHIBI'IING EFFECT OF CALCIUM gggilgYL-Z LACTYLATE ON PARTIALLY BAKED DINNE R Time in Days for the Development of- Package Identification First Multiple Complete Colony Colonies Coverage Control 10 12 14 1% Calcium Propionate 19 24 No mold development after 30 days 1% Calcium capryl-2 lactylate N o mold development after 30 days 2% Calcium capry12 lactylate- Example .The inhibition of mold growth on leather with calcium capryl-2 lactylate Five strips of leather approximately one by three inches, were soaked and washed in a detergent, rinsed and dried. The stripswere dipped in the following solutions:

(1) Ethanol (control) (2) 1% calcium capryI-Zlactylate in ethanol (3) 2.5% calcium capryl-2 lactylate in ethanol (4) 1% benzoic acid in ethanol (5) 2.5% benzoic acid in ethanol The excess solution was blotted from the strips with.

filter paper. The leather was suspended from a wire in the room for twenty-four hours to allow the ethanol to evaporate and. to inoculate the leather with air borne spores. The strips were incubated by hanging them in a large jar. .Water was placed in the bottom of the jar to insure high humidity during the storage period.

The leather was examined periodically for evidence of Example 6.-Tests studying the action of calcium capryl-2 lactylates on skin fungi Three people voluntarily served as test subjects to determine the eifectiveness of calcium capryl-2 lactylate in treating cases of skin fungi. One case was a fungus in- Eection in the auditory canal, the other two were athletes foot.

a For treatment of these infections an ointment and a powder were prepared. The ointment consisted of :alcium capryl-2,1actylate dissolved. in petroleum jelly formed by melting'the two materials together and stirring until cool.

was evaporated and the powder. passed through a 60 mesh screen to break up lumps. The finished powder contained 2.5 calcium capryl-2 lactylate by weight.

The ointment was used in the treatment of the infection in the auditory canal and the powder was used in the treatment of athletes foot. In each instance after single daily applications for one week complete symptomatic relief was obtained. 7

We claim:

1. A method for the protection of loci susceptible to the growth of undesired microorganisms against such growth which comprises subjecting such loci to an effec- The powder was formed by preparing an ethanol solution of the lactylate and stirring this solution into. talcum powder until a paste was formed. The ethanol.

is a number from 1 to n.

tive amount of an acylated boxylic acid of the formulawherein RC0 is an acyl radical of a fatty acid of from 8 to 12 carbon atoms, each of A and B are selected from the group consisting of hydrogen and methyl, Y is a cation, x as an average is a number-between about.0.3 and i 6, n is an integer equal to the'valence of the cation Y and I m is'a number from 1 to n. a

2. A method for the protection of loci susceptible to the growthof .,undesired microorganisms: against such growth which comprises subjecting such loci to an effective amount of an acyla'ted product of an .a-hYdI'OXY carboxylic acid of the formula (13H: Roomc1 10o).o ..,Y oH)..-m wherein RCO'is an acyl radical of a fatty acid of from 8 to 12 carbon atoms, .Y is a cation, x as an average is a number between 0.3 and 3, n is an integer equal to the valence of the cation Yand m is a number from 1 to n.

V 3. A method for the protection of loci susceptible to. the growth of undesired microorganisms against such growth which comprises subjecting such loci to an eifective amount of an acylated product of an a-hydroxy carboxylic acid of the formula wherein Y is a cation, x is a number between 1 and 3, n-

is an integer equal to the valence of the cation Y and m 4. A method for the protection of loci, susceptible to the growth of undesired microorganisms against such.

growth which comprises subjecting such loci to an effective amount of an alkali metal capryl lactylate containing an average of l to 3 lactylic groups.

5. A method for the protection of locisusceptible to the growth of undesired microorganisms against such growth which comprises subjecting such loci to an edective amount of an alkaline earth metal capryl lactylate.

containing an average of l to 3 lactylic groups; 6. A method for the protection of locisusceptible growth which comprisessubjecting such loci to an effective amount of capryl lactylic acid containing an average I of 1 to 3 lactylic groups.

7. A method for the protection of locisusceptible to the growth of undesired microorganisms against such growth which comprises subjecting such loci-to an effec-. tive amount of calcium capryl lactylate containing an. average of 1 to 3 lactylic groups.

8. A method for the protection of loci susceptible to the growth of undesired microorganisms against such growth which comprises subjecting such loci to an eifec-. tive amount of calcium capryl-2 lactylate.

9. A method for theprotection of foodstutf susceptible to the growth of undesired microorganisms against such growth which comprises subjecting such foodstufi to an. effective amount of a non-toxic acylated product ofan r a-hydroxy carboxylic acid of the formula I A t i [nooior oonmmYtomncn wherein RC0 is an acyl radical of a fatty acid of from 8 7 ot 12 carbon atoms, each of A and B are selected from the group consisting of hydrogen and methyl, Y is a cation, 1: as an average is a number between 0.3 and 6, n is an integer equal to the valence of the cationY and m is a number from 1 to n.

product of an a-hydroxy carthe growth of undesired microorganisms against such 9 10. A method for the protection of foodstuff susceptible to the growth of undesired microorganisms against such growth which comprises incorporating in such foodstuff an effective amount of a non-toxic acyla-ted product of an a-hydroxy carboxylic acid of the formula wherein RC is an acyl radical of a fatty acid of from 8 to 12 carbon atoms, each of A and B are selected from the group consisting of hydrogen and methyl, Y is a cation, x as an average is a number between 0.3 and 6, n is an integer equal to the valence of the cation Y and m is a number from 1 to n.

11. A method for the protection of foodstuff susceptible to the growth of undesired microorganisms against such growth which comprises coating such foodstuff with an effective amount of a non-toxic acylated product of an a-hydroxy carboxylic acid of the formula wherein RC0 is an acyl radical of a fatty acid of from 8 to 12 carbon atoms, each of A and B are selected from the group consisting of hydrogen and methyl, Y is a cation, x as an average is a number between 0.3 and 6, n is an integer equal to the valence of the cation Y and m is a number from 1 to n.

12. A method for the protection of foodstuff susceptible to the growth of undesired microorganisms against such 10 growth which comprises wrapping such foodstuff in a sheet of material carrying uniformly distributed over the area thereof an effective amount of a non-toxic acylated product of an a-hydroxy carboxylic acid of the formula wherein RC0 is an acyl radical of a fatty acid of from 8 to 12 carbon atoms, each of A and B are selected from the group consisting of hydrogen and methyl, Y is a cation, x as an average is a number between 0.3 and 6, n is an integer equal to the valence of the cation Y and m is a number from 1 to n.

References Cited by the Examiner LEWIS GOTTS, Primary Examiner.

S; K. ROSE, Assistant Examiner. 

1. A METHOD FOR THE PROTECTION OF LOCI SUSCEPTIBLE TO THE GROWTH OF UNDESIRED MICROORGANISMS AGAINST SUCH GROWTH WHICH COMPRISES SUBJECTING SUCH LOCI TO AN EFFECTIVE AMOUNT OF AN ACYLATED PRODUCT OF AN A-HYDROXY CARBOXYLIC ACID OF THE FORMULA 